The pyruvate dehydrogenase complex (PDHC) will be isolated from human kidney, using techniques which have been well standardized for tissues of other mammals. PDHC will also be isolated from kidney of a patient who died with well documented PDHC deficiency. PDHC from the patient and from controls will be compared by slab gel electrophoresis and by kinetic techniques (by comparing Km for substrates, Ki for inhibitors, susceptibility to heat inactivation, and dependence on ionic strength and pH). PDHC peptides will also be compared by slab gel electrophoresis in cultured skin fibroblasts from PDHC deficient patients and controls, using purified PDHC as a marker to identify the positions of the four main PDHC peptides. The cell lines studied will include a line of cultured fibroblasts from the deceased patient from whose tissue PDHC has been isolated. They will also include cells from nineteen other patients, in whose cells PDHC-deficiency has been demonstrated by direct assay. These include patients in whom there is kinetic evidence for deficiencies in each of the three major enzymes of the PDHC complex. These experiments will demonstrate whether or not electrophoretic abnormalities in component peptides of PDHC occur in all, in some, or in none of the patients with PDHC-deficiency. They will provide the first direct evidence, on the level of the proteins, for or against the existence of structural mutations affecting the peptides of PDHC in humans, whose tissues have been found deficient in PDHC activity.